中国滇西地区葡萄炭疽病病原菌的鉴定及其杀菌剂筛选  

Identification of the Pathogens Causing Grape Anthracnose in West Yunnan,China and Selection of Effective Fungicides

作　　者：苏奇倩 何鹏搏 何俊 张华玲 周会明 冯之俊 谢艳兰 苏向宇 于龙凤 谭万忠 SU Qiqian;HE Pengbo;HE Jun;ZHANG Hualing;ZHOU Huiming;FENG Zhijun;XIE Yanlan;SU Xiangyu;YU Longfeng;TAN Wanzhong(College of Biotechnology and Engineering,West Yunnan University,Lincang Yunnan 677000,China;College of Plant Protection,Yunnan Agricultural University,Kunming 650000,China;Yunnan Black Tea Group Co.Ltd.,Fengqing Yunnan 675900,China)

机构地区：[1]滇西科技师范学院生物工程技术学院,云南临沧677000 [2]云南农业大学植物保护学院,昆明650000 [3]中国云南省临沧市凤庆县滇红茶集团,云南凤庆675900

出　　处：《西南大学学报（自然科学版）》2024年第8期31-44,共14页Journal of Southwest University(Natural Science Edition)

基　　金：国家自然科学基金地方专项(32360411);中央引导地方科技发展资金项目(202207AB110007);云南省科技科技人才和平台计划建设(2018DH011);地方高校联合专项(202001BA070001-187);云南省临沧市科学技术具科技创新团队建设项目(202204AC100001-TD01)

摘　　要：本研究旨在弄清滇西地区葡萄炭疽病的病原菌及筛选有效抑制病原菌的杀菌剂.从云南西部不同地区采集葡萄病叶样品,通过分离培养和纯化共获得13株真菌纯培养,经离体叶片接种证病试验验证,其中7株刺盘孢真菌(编码PT23-01~PT23-07)为葡萄炭疽病的致病菌.通过PCR扩增获得这7个致病菌株的内转录间区(ITS)、肌动蛋白、钙调蛋白、组蛋白3和β-微管蛋白2基因(ACT,CAL,HIS3,TUB2)序列并提交到NCBI GenBank,用Megablast软件在线进行Blast-n比对结果显示,这7个菌株与刺盘孢属相应种类的相似性分别达到99.27%~100%.进一步用MEGA7.0构建基于4个蛋白基因的系统演化树分析,其显示PT23-01,PT23-03,PT23-04和PT23-05与盘长孢刺盘孢(Colletotrichum gloeosporioides jo.3)聚于同一末端分枝(BT=100),PT23-2和PT23-07与果生刺盘孢(C.fructicola AV103)处于同一分枝(BT=94),而PT23-06则与暹罗刺盘孢(C.siemense ABS125375)聚为同一分枝(BT=98),这与Blast-n比对的结果一致.在所鉴定的3种病菌中,C.gloeosporioides是优势病原菌,从滇西各地区样品中均被分离到;而C.siemense在文献中尚未见有侵染葡萄的报道,是导致葡萄炭疽病的一种新病原菌;本研究简要地描述了这3种病菌的形态特征.通过菌落生长法测定,8种供试杀菌剂都能不同程度地抑制盘长孢刺盘孢的菌落生长,不同供试药剂对同一种菌的抑菌效果没有显著差异(p>0.05).25%甲硫乙唑醇SA和30%苯甲吡唑脂SA的抑制作用最强,抑菌率分别达92.25%和95.87%(p<0.01),EC 50值分别为1.103 mg/mL和1.521 mg/mL.室内测定结果显示,两种药剂对葡萄炭疽病也具有很好的控制效果,控病率分别达到85.03%和91.54%(p<0.01).本研究结果为滇西地区葡萄炭疽病的正确诊断鉴定和有效控制奠定了科学基础.The aim of present study was to identify the pathogens causing grapevine anthracnose in west Yunnan(Diqing,Baoshan,Lincang and Puer)and to select effectivefungicides forcontrol of the disease.Through isolation and purification,13 fungal isolates were obtained and 7 isolates(coded as PT23-01 to PT23-07)were verified as pathogens of grapevineanthracnose after pathogenicity tests following Koch s Postulates.The internal transcribed spacer(ITS),Actin,Calmodulin,Histone 3 andβ-Tubulin genes(ATC,CAL,HIS3,TUB2)of the 7 pathogenic isolates were PCR-amplified,sequenced and submitted to NCBI GenBank.The results of Blast-n alignment of these genes using the Megablast software showed that all the isolates were highly identical with corresponding Colletotrichum spp.(Per.Ident.>99.27%).The neighbor-joining tree based on concatenated sequence of the 4 protein genes was constructed using MEGA 7.0 software.T23-01,PT23-03,PT23-04 and PT23-05 were clustered with Colletotrichum gloeosporioides jo.3(BT support=100),PT23-2 and PT23-07 were with C.fructicola AV103(BT support=94),and PT23-06 was with C.siemense ABS125375(BT support=98).Among the 3 species,C.gloeosporioides is the most dominant species distributed all over west Yunnan and C.siamense is a newly recorded pathogen of grapevine anthracnose.The morphologies of three fungal species were described in this paper.Eight tested fungicides showed different degree of inhibitory effect on the colony growth of Colletotrichum spp.,and there was no significant difference in the fungicidal effect of different fungicides on the same fungus(p>0.05).The 25%Methanethiolacetate SA and 30%Benzyl pyrazolid SA had the highest inhibitory effect on fungal growth,with inhibitive rate of 92.25%and 95.87%(p<0.01),respectively.The toxicity regression functions of the two fungicides were established and their median effective concentration(EC 50)calculated from the functions was 1.103 mg/mL and 1.521 mg/mL,respectively,indicating that both fungicides were very effective against the anthracnose pathogen

关 键 词：葡萄炭疽病 多基因鉴定 刺盘孢菌 杀菌剂筛选 抑菌控病效果 

分 类 号：S436.631[农业科学—农业昆虫与害虫防治]